Cefuroxime is widely used to prevent postoperative endophthalmitis after cataract surgery. For its potential applications in vitreoretinal surgeries, we previously reported its cytotoxicity on human retinal pigment epithelium (RPE) cells. This study further elucidated its mechanisms of cell death.

On the cell viability assay, exposure to 0.25 mg/mL cefuroxime for 24 h or 25 mg/mL for 1 h impaired mitochondrial functions and damaged 45% and 23%, respectively, of cells. Morphologically, trypan blue-stained dead and shrunk cells were observed in dose and time relationships. On the ultrastructure, exposure to 2.5 mg/mL cefuroxime for 24 h lead to swollen organelles, and 7.5-250 mg/mL cefuroxime induced more swelling and disorganization of organelles, disruption of cell membranes, and even cell lysis. On the mechanisms of cell death, morphological staining and quantitative flow cytometry consistently showed more apoptosis (annexin V-FITC) than necrosis (propidium iodide) at dose- and time-dependent manners. On the TUNEL staining, exposure to 7.5-250 mg/mL cefuroxime caused late apoptotic DNA fragmentation. Apoptosis was reduced, but not eliminated, by inhibitors of caspase-1, -2, -3 or -9; apoptosis was induced by addition of these caspase activators. Caspases-6 and -8 did not affect cefuroxime-induced apoptosis. Cyclosporine A stabilized mitochondria and decreased apoptosis in a dose relationship. Lysosomes did not play a major role in cefuroxime-induced apoptosis.

Conclusions

The mechanisms of cefuroxime cytotoxicity in human RPE cells are majorly apoptosis and somewhat necrosis at time- and dose-dependent manners. Apoptotic cell deaths include a caspase-dependent pathway via caspase-2, mitochondria and caspase-3, and a caspase-independent pathway. Ultrastructural damage and mitochondria dysfunction are noted within 24 hours. We conclude that 2.5 mg/mL cefuroxime damages RPE cells and recommend safety doses in vitreoretinal surgeries.